DOI:10.1002/chem.201602195 Full Paper & Prebiotic Chemistry |Hot Paper| The Role of Aqueous Aerosols in the “Glyoxylate Scenario”: An Experimental Approach Margarita R. Marín-Yaseli, ElenaGonzµlez-Toril,Cristina Mompeµn,and Marta Ruiz- Bermejo*[a] Abstract: The origin of life is one of the fundamentalques- origin of life. The soluble andinsoluble HCN polymers syn- tions in science. Eschenmoser proposed the “glyoxylate sce- thetizedwere analyzed by GC-MS. We identified, for the first nario”, in whichplausible abiotic synthesis pathways were time, glyoxylic acid in thesepolymers, together with some suggested to be compatible with the constraints of prebiotic constituents of the reductive tricarboxylic acid cycle, amino chemistry.Inthis proposal,the stem compound is HCN. In acids and several N-heterocycles. The findings presented this work, we explore the “glyoxylate scenario” through sev- herein, asthe first globalapproachtothe “glyoxylate scenar- eral syntheses of HCN polymers, paying particularattention io”, give full effect to this hypothesis and prove that aque- to the role of the aqueous aerosols,together with statistical ous aerosols could play an important role in this plausible methods, as astep to elucidate the synthetic problem of the scene of the origin of life. Introduction organics,[6] and they are considered key in the primeval evolu- tion of protometabolism and informational systems.[7] Biogenesis can be understood as apuzzling and intriguing ret- On the other hand, the possible importance of aerosols in rosynthetic problem. One of the main challenges in solving the origin of life on Archean Earth has been emphasized in this great question of currentscience is that there are awide recentyears, and we couldassume that the bubble-aerosol- variety of uncertainties about the geochemistryofthe ancient droplet cycle (the bursting of bubbles that injectsinto the at- Earth. In this context, Eschenmoser proposed the “glyoxylate mosphere the aerosol particles and their subsequent conden- scenario”, in which plausible abioticsynthesis pathways are sation as droplets) was active in the Archean epoch. The aque- suggested to be compatible with the constraintofprebiotic ous aerosols can be considered as “prebiotic microreactors”[8] chemistry.[1,2] In this hypothetical prebiotic scenario, glyoxylate and show an efficient variation in the reactivity of the sys- and its formal dimer,dihydroxyfumarate, are suggestedtobe tems.[9] They can enhancethe yield of polar organic com- the key startingmaterials of the chemical constitution of apos- pounds,[10] improvethe formationofdeterminate organics sible metabolism, serving as asource of the main biomono- against others,[7a] and also influx positivelyinnon-enzymatic mers, such as sugars, amino acids, pyrimidines and the constit- polymerization reactions.[11] In addition, currently there is an in- uents of the reductive tricarboxylic acid cycle (rTCAC, creasinginterest in the reactivity of organicsatthe air–water Scheme 1). In the “glyoxylate scenario”, the stem compound is interfaces.[12] HCN, aubiquitous molecule in the universe, and although sev- Recently,the use of areactionmatrix has been suggested eral plausible prebiotic sources of glyoxylate have been consid- for screening prebioticreactions to test the conditions that ered,[3–5] there are no examples that have been previously re- could likely lead to the emergence of aprimeval biochem- ported in the literature that prove the production of glyoxylate istry.[13] Along this line of thinking, experimental studies have from HCN, as originally suggestedbyEschenmoser.Indeed, in been carried out to determinate the plausible processes that this same context,Eschenmoser suggestedahypothetical rela- may lead to the symmetry breaking,using chiral inorganic sub- tionship between the HCN andthe constituents of the rTCAC strates as model standards and statisticalcalculations.[14] In this (Scheme 2). Additionally,the named HCN polymers have great work, following these proposals, we suggest an experimental relevance in studies on the primeval synthesis of the first bio- approachtoevaluatingthe “glyoxylate scenario” through sev- eral syntheses of HCN polymers, payingparticular attention to the role of the aqueous aerosols, togetherwith statistical [a] M. R. Marín-Yaseli, Dr.E.Gonzµlez-Toril, C. Mompeµn, Dr.M.Ruiz-Bermejo methods, as astep to elucidating the synthetic problem of the Departamento de Evolución Molecular origin of life. Thus, we assayed the production of HCN poly- Centro de Astrobiología (INTA-CSIC), Ctra. Torrejón-Ajlavir mers using differentexperimental conditions to evaluatethe km 4,8, 28850 Torrejón de Ardoz, Madrid (Spain) E-mail:[email protected] effects of the environmental variables, such as aqueous aero- Supportinginformation for this article can be found under sols, reaction time, presence of ammonium ion and oxygen http://dx.doi.org/10.1002/chem.201602195. and salinity,onthe yield of the polymerizationprocess itself as Chem. Eur.J.2016, 22,12785 –12799 12785 2016 Wiley-VCH Verlag GmbH &Co. KGaA, Weinheim Full Paper Scheme1.Reductivetricarboxylic acid cycle (rTCAC; adapted from Guzman and Martin). This cyclehas been proposed as aplausible mechanism for carbon fixationand energy storage at the time of emergence of life. This cycle is the central axis of the universal metabolism. The combination of rTCAC and its exit products operates as afactoryfor the synthesis on the main classes of biomolecules. The components of this cycleidentified in HCN polymers are indicated in boxes. Scheme2.Hypothetical relationships between HCN oligomersand constituents of the rTCAC (adaptedfrom Eschenmoser 2007).[1] The carboxylic acids that are constituents of the rTCAC are indicated in boxes. [a] C4-diacid tautomers =2-hydroxyoxaloacetic acid,2,3-dihydroxyfumaric acid and 2,3-dihydroxymaleic acid. The glyoxylic acid (c9), pyruvic acid (c10), alanine (a3), asparticacid (a7)and glutamic acid (a13), were identifiedinHCN polymers by GC-MS. Chem. Eur.J.2016, 22,12785 –12799 www.chemeurj.org 12786 2016 Wiley-VCH Verlag GmbH &Co. KGaA, Weinheim Full Paper well as in the monomeric diversity present in the HCN poly- In this work, we used acid hydrolysis conditions because mers synthetized. Statistical calculations and representations of they are astandard method to screening for awide variety of the results helpedustointerpretall data generated in simple polar compounds of biological interestpresentincomplex overviewsand to relate the different environmental variables substances produced under possible prebiotic conditions, such with the synthetic and analytical results to see what experi- as tholins and HCN polymers, and are also present in meteor- mental conditions are the most favorable from the point of ites.[18,19] All the hydrolyzed samples were derivatized with view of the “glyoxylate scenario”. To our knowledge, there BSTFAtoobtain the correspondingTMS derivatives of the have been only two works on the production of organic mole- monomers indicated above.This derivatization method is not cules under possible prebiotic conditions that directly take specific for each type of compound mentioned, but for compa- into consideration the “glyoxylate scenario” as it was proposed rative purposes,itprovidesanexcellent general overview of by Eschenmoser.[15,16] In these works, the startingreactants are the polar molecules present in all the HCN polymers synthe- glyoxylate and dihydroxyfumarate. Herein, the starting reac- tized. Therefore, this analytical methodology waschosen be- tants are NH4CN or NaCN. cause it allowed us to discriminate among the various synthe- All the HCN polymers synthetized herein (using synthetic sis conditions tested to determine which of them are the most conditions shown in Table 1) were qualitativelyanalyzed for favorablefrom the point of view of the “glyoxylate scenario”. polar monomers (mainly amino acids, carboxylic acids and N- Additionally,monomers never yet identified in HCN polymers heterocycles) by GC-MS, after acid hydrolysis. It is well-known were detected under the experimental conditions assayed that the hydrolysis conditions have anotable influence on the herein.Notably,wereport for the first time the identification detection, identification and quantification of the monomers of glyoxylic acid (c9), in accordance with the originalproposal present in the HCN polymers.[6a,7a, 17] of Eschenmoser.All the monomers identified are summarized Table 1. Experimental conditionsused to produce the HCN polymers. The averages and the standard deviations of the amounts of the insoluble HCN polymers and the final pH of the reaction suspensions were calculated by using the data of at leastfour independent syntheses. [a] [b] [c] [d] Sample Reactant t [d] Aerosol Salts O2 Weight [mg] Final pH %inweight 1NH CN 3 + 15.3 3.0 9.40 0.03 4.37 4 ÀÀ Æ Æ 2NH CN 3 + + 5.6 1.6 9.53 0.03 2.81 4 À Æ Æ 3NH CN 3 9.4 0.8 9.62 0.09 5.34 4 ÀÀÀ Æ Æ 4NH CN 3 + 1.1 0.3 9.63 0.13 2.16 4 ÀÀ Æ Æ 5NH CN 3 ++ 29.5 2.7 9.64 0.02 30.55 4 À Æ Æ 6NH CN 3 +++ 10.2 2.2 9.56 0.07 14.36 4 Æ Æ 7NH CN 3 + 19.6 2.2 9.65 0.02 20.48 4 À À Æ Æ 8NH CN 3 ++ 8.1 2.3 9.60 0.05 8.79 4 À Æ Æ 9NH CN 30 + 62.0 3.1 9.90 0.02 19.34 4 ÀÀ Æ Æ 10 NH CN 30 + + 8.0 1.4 9.20 0.23 1.25 4 À Æ Æ 11 NH CN 30 28.8 0.8 9.91 0.04 6.76 4 ÀÀÀ Æ Æ 12 NH CN 30 + 2.1 1.3 9.10 1.00 1.30 4 ÀÀ Æ Æ 13 NH CN 30 ++ 36.4 2.5 9.87 0.02 21.6 4 À Æ Æ 14 NH CN 30 +++ 23.2 9.3 8.63 0.64 9.46 4 Æ Æ 15 NH CN 30 + 41.4 1.3 9.91 0.02 14.28 4 À À Æ Æ 16 NH CN 30 ++ 4.4 3.2 9.44 0.09 11.35 4 À Æ Æ 17 NaCN